High-Throughput Kinetic Screening of Hybridomas to Identify High-Affinity Antibodies Using Bio-Layer Interferometry
In this study, we have applied Bio-Layer Interferometry to screen hybridoma clones based on disassociation rates using the OctetRED 384 platform. Using the OctetRED platform, we were able to screen 2000 clones within 24 hours and select clones containing high-affinity antibodies for further expansion and subsequent characterization. Using this method, we were able to identify several clones producing high-affinity antibodies that were missed by ELISA. (Source: Journal of Biomolecular Screening)
Source: Journal of Biomolecular Screening - March 24, 2015 Category: Molecular Biology Authors: Lad, L., Clancy, S., Kovalenko, M., Liu, C., Hui, T., Smith, V., Pagratis, N. Tags: Original Research Source Type: research
A Fully Automated Primary Screening System for the Discovery of Therapeutic Antibodies Directly from B Cells
This article will describe the design, construction, and commissioning of a bespoke automated screening platform and two examples of how it was used to screen for antibodies against two targets. (Source: Journal of Biomolecular Screening)
Source: Journal of Biomolecular Screening - March 24, 2015 Category: Molecular Biology Authors: Tickle, S., Howells, L., O'Dowd, V., Starkie, D., Whale, K., Saunders, M., Lee, D., Lightwood, D. Tags: Original Research Source Type: research
How to Successfully Patent Therapeutic Antibodies
Therapeutic antibodies have become an established class of drugs for the treatment of a variety of diseases, especially cancer and autoimmune/inflammatory disorders, and a sufficient patent protection is a prerequisite for their successful commercialization. As monoclonal antibodies and their therapeutic potential have been well known for decades, the mere production of yet another therapeutic antibody is in many jurisdictions not considered a patentable invention. In contrast, antibodies with novel structural features and/or improved properties may be patentable. When drafting the claims, care should be taken to obtain a ...
Source: Journal of Biomolecular Screening - March 24, 2015 Category: Molecular Biology Authors: Lahrtz, F. Tags: Review Articles Source Type: research
Accelerated Formulation Development of Monoclonal Antibodies (mAbs) and mAb-Based Modalities: Review of Methods and Tools
This article reviews the latest advances in methods of formulation screening, which allow not only the high-throughput selection of the most suitable formulation but also the prediction of stability properties under manufacturing and long-term storage conditions. We describe how the combination of automation technologies and high-throughput assays creates the opportunity to streamline the formulation development process starting from early preformulation screening through to commercial formulation development. The application of quality by design (QbD) concepts and modern statistical tools are also shown here to be very ef...
Source: Journal of Biomolecular Screening - March 24, 2015 Category: Molecular Biology Authors: Razinkov, V. I., Treuheit, M. J., Becker, G. W. Tags: Review Articles Source Type: research
Discovery of Functional Antibodies Targeting Ion Channels
Ion channels play critical roles in physiology and disease by modulation of cellular functions such as electrical excitability, secretion, cell migration, and gene transcription. Ion channels represent an important target class for drug discovery that has been largely addressed, to date, using small-molecule approaches. A significant opportunity exists to target these channels with antibodies and alternative formats of biologics. Antibodies display high specificity and affinity for their target antigen, and they have the potential to target ion channels very selectively. Nevertheless, isolating antibodies to this target cl...
Source: Journal of Biomolecular Screening - March 24, 2015 Category: Molecular Biology Authors: Wilkinson, T. C. I., Gardener, M. J., Williams, W. A. Tags: Review Articles Source Type: research
New Horizons in Therapeutic Antibody Discovery: Opportunities and Challenges versus Small-Molecule Therapeutics
Antibody drugs have become an increasingly significant component of the therapeutic landscape. Their success has been driven by some of their unique properties, in particular their very high specificity and selectivity, in contrast to the off-target liabilities of small molecules (SMs). Antibodies can bring additional functionality to the table with their ability to interact with the immune system, and this can be further manipulated with advances in antibody engineering. This review summarizes what antibody therapeutics have achieved to date and what opportunities and challenges lie ahead. The target landscape for large m...
Source: Journal of Biomolecular Screening - March 24, 2015 Category: Molecular Biology Authors: Smith, A. J. Tags: Review Articles Source Type: research
JBS Special Issue on Therapeutic Antibody Discovery and Development: Biologics Come of Age
(Source: Journal of Biomolecular Screening)
Source: Journal of Biomolecular Screening - March 24, 2015 Category: Molecular Biology Authors: McGivern, J. G., Howes, R. Tags: From the Guest Editors Source Type: research
The Acute Extracellular Flux (XF) Assay to Assess Compound Effects on Mitochondrial Function
Numerous investigations have linked mitochondrial dysfunction to adverse health outcomes and drug-induced toxicity. The pharmaceutical industry is challenged with identifying mitochondrial liabilities earlier in drug development and thereby reducing late-stage attrition. Consequently, there is a demand for reliable, higher-throughput screening methods for assessing the impact of drug candidates on mitochondrial function. The extracellular flux (XF) assay described here is a plate-based method in which galactose-conditioned HepG2 cells were acutely exposed to test compounds, then real-time changes in the oxygen consumption ...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Wang, R., Novick, S. J., Mangum, J. B., Queen, K., Ferrick, D. A., Rogers, G. W., Stimmel, J. B. Tags: Technical Notes Source Type: research
Automation of a Phospho-STAT5 Staining Procedure for Flow Cytometry for Application in Drug Discovery
Drug discovery often requires the screening of compound libraries on tissue cultured cells. Some major targets in drug discovery belong to signal transduction pathways, and PerFix EXPOSE* allows easy flow cytometry phospho assays. We thus investigated the possibility to further simplify and automate this assay, to allow the direct screening of drugs targeting signaling pathways. We show here the sensitivity of this fully automated assay on human growth hormone (hGH)-driven JAK/STAT5-activated IM-9 cells, and we discuss the throughput of this system, which is compatible with medium-throughput drug screening. Because the kit...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Malergue, F., van Agthoven, A., Scifo, C., Egan, D., Strous, G. J. Tags: Technical Notes Source Type: research
Using the BioAssay Ontology for Analyzing High-Throughput Screening Data
High-throughput screening (HTS) is the main starting point for hit identification in drug discovery programs. This has led to a rapid increase of available screening data both within pharmaceutical companies and the public domain. We have used the BioAssay Ontology (BAO) 2.0 for assay annotation within AstraZeneca to enable comparison with external HTS methods. The annotated assays have been analyzed to identify technology gaps, evaluate new methods, verify active hits, and compare compound activity between in-house and PubChem assays. As an example, the binding of a fluorescent ligand to formyl peptide receptor 1 (FPR1, i...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Zander Balderud, L., Murray, D., Larsson, N., Vempati, U., Schurer, S. C., Bjareland, M., Engkvist, O. Tags: Original Research Source Type: research
Open Access to High-Content Clonogenic Analysis
This article describes an experimental and multiparametric image analysis workflow for clonogenic assays in multiwell format, named the Colony Assay Toolbox (CAT). CAT incorporates a cellular-level resolution of individual colonies and facilitates the extraction of phenotypic information, including the number and size of colonies and nuclei, as well as morphological parameters associated with each structure. Furthermore, the pipeline is capable of discriminating between colonies composed of senescent and nonsenescent cells. We demonstrate the accuracy and flexibility of CAT by interrogating the effects of 2 preclinical com...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Ricci, F., Subramanian, A., Wade, M. Tags: Original Research Source Type: research
High-Content Phenotypic Screening and Triaging Strategy to Identify Small Molecules Driving Oligodendrocyte Progenitor Cell Differentiation
We describe the details of an automated, cell-based, morphometric-based, high-content screen that is used to identify small molecules eliciting the differentiation of OPCs after 3 days. Primary screening was performed using rat CG-4 cells maintained in culture conditions that normally support a progenitor cell–like state. From a library of 73,000 diverse small molecules within the Sanofi collection, 342 compounds were identified that increased OPC morphological complexity as an indicator of oligodendrocyte maturation. Subsequent to the primary high-content screen, a suite of cellular assays was established that ident...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Peppard, J. V., Rugg, C. A., Smicker, M. A., Powers, E., Harnish, E., Prisco, J., Cirovic, D., Wright, P. S., August, P. R., Chandross, K. J. Tags: Original Research Source Type: research
Detection of Cell Aggregation and Altered Cell Viability by Automated Label-Free Video Microscopy: A Promising Alternative to Endpoint Viability Assays in High-Throughput Screening
Automated phase-contrast video microscopy now makes it feasible to monitor a high-throughput (HT) screening experiment in a 384-well microtiter plate format by collecting one time-lapse video per well. Being a very cost-effective and label-free monitoring method, its potential as an alternative to cell viability assays was evaluated. Three simple morphology feature extraction and comparison algorithms were developed and implemented for analysis of differentially time-evolving morphologies (DTEMs) monitored in phase-contrast microscopy videos. The most promising layout, pixel histogram hierarchy comparison (PHHC), was able ...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Aftab, O., Fryknas, M., Hammerling, U., Larsson, R., Gustafsson, M. G. Tags: Original Research Source Type: research
A High-Throughput Phenotypic Screen of Cytotoxic T Lymphocyte Lytic Granule Exocytosis Reveals Candidate Immunosuppressants
We screened the National Institutes of Health’s Molecular Libraries Small Molecule Repository for inhibitors of cytotoxic T lymphocyte (CTL) lytic granule exocytosis by measuring binding of an antibody in the extracellular solution to a lysosomal membrane protein (LAMP-1) that is transferred to the plasma membrane by exocytosis. We used TALL-104 human leukemic CTLs stimulated with soluble chemicals. Using high-throughput cluster cytometry to screen 364,202 compounds in a 1536-well plate format, we identified 2404 initial hits: 161 were confirmed on retesting, and dose–response measurements were performed. Seven...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Zhao, Z., Haynes, M. K., Ursu, O., Edwards, B. S., Sklar, L. A., Zweifach, A. Tags: Original Research Source Type: research
Tetracycline-Based System for Controlled Inducible Expression of Group III Metabotropic Glutamate Receptors
A stable and inducible expression of metabotropic glutamate receptor type 4, 7, and 8 was obtained in T-REx 293 cells using the tetracycline system. Tetracycline administration to the cell medium resulted in rapid induction and time-dependent expression of mGlu receptors, which also correlates with its functionality in a cAMP accumulation assay. The pharmacological properties of recombinant mGlu receptors were verified using orthosteric and allosteric ligands. Data suggest that the Tet-on inducible system is suitable for functional mGlu receptors’ expression and characterization by means of the cAMP accumulation assa...
Source: Journal of Biomolecular Screening - February 19, 2015 Category: Molecular Biology Authors: Chruścicka, B., Burnat, G., Brałski, P., Chorobik, P., Lenda, T., Marciniak, M., Pilc, A. Tags: Original Research Source Type: research
